Genes encoding phosphinothricin-N-acetyl-transferase (PAT), EC 2.3.1.183, are routinely used as selectable markers in transgenic events in plants and were originally isolated from the common aerobic soil actinomycete, Streptomyces viridochromogenes. The PAT enzyme catalyzes the acetylation of phosphinothricin, detoxifying it into an inactive compound resulting in the accumulation of ammonia and cell death (Murakami T., Anzai H., Imai S., Satoh A., Nagaoka K., Thompson C. J. (1986). Mol Gen Genet 205:42-50; Twell D., Klein T. M., Fromm M. E., McCormick S. (1989). Plant Physiol 91:1270-1274.) . Transformed plant cells expressing PAT can therefore be selected using glufosinate.
Companies who develop and market recombinant DNA traits for planting seed products formulate, implement and adhere to strict product stewardship plans. These stewardship plans require the use of validated quantitative and qualitative protein detection methods for various components of the recombinant trait to track trait introgression and seed production activities, as well as monitoring grain harvest. These detection methods must be facile and robust enough to use under GLP and non-GLP conditions. Moreover the methods must be user friendly enough to be easily employed by farmers in the field, grain dealers at the silo, and customs officials at the borders. Therefore, robust, high quality, user friendly protein detection methods and commercial kits are useful and necessary.
While ELISAs are well known in the art, developing robust, high quality, validated ELISA methods that are reproducibly able to detect a particular transgenic product in an array of plant tissue in both lab and field settings is neither trivial nor routine. Still more challenging is to find antibody pairs that are particularly suited to the development of a lateral flow strip and/or ELISA for detecting a functioning PAT transgenic event.